Prochlorococcus, the dominant photosynthetic organism throughout the oligotrophic oceans, serves as a model for microbial oceanography, linking information from the genomic level to biogeochemical cycles. Most physiological research has focused on understanding differences between High Light (HL)-adapted and Low Light (LL)-adapted Prochlorococcus ecotypes (Moore and Chisholm 1999), or distinguishing differences among the HLI and HLII Prochlorococcus subclades (Johnson et al 2006). However the LL-adapted Prochlorococcus subclades are more genetically diverse, but understanding physiological differences between the LL clades has been difficult because only a few cultured representatives existed as of 2008. We employed a high throughput culturing method to isolate new Prochlorococcus representatives from LL clades I-IV, as determined by sequence analysis of the 16S-23S rRNA ITS region. We used these new isolates along with more established isolates to examine a variety of ecophysiological parameters in order to determine possible defining phenomic characteristics that contribute to the ecology and evolution of the phylogenetically-distinct LL subclades. Based on studies of natural populations in the field (Mahlstrom et al 2010), the LLI ecotype exhibited depth distributions at Bermuda Atlantic Time Series station that were 'intermediate' between HL ecotypes and the other LL ecotypes. Our physiological studies of 4 LLI strains support this hypothesis. The LL ecotype strains exhibit light-dependent and temperature-dependent growth responses more similar to HL strains, but exhibit pigment content similar to other LL ecotypes, such as higher chlorophyll b to chlorophyll a ratio and measurable phycoerythrin pigments. The LLIV ecotype isolates consistently exhibit more extreme photoinhibition of growth than other LL-adapted ecotypes, some which cannot grow at irradiances as low as 50 mmol quanta m-2 s-1. The LLIV ecotype isolates also exhibit higher chlorophyll and phycoerythrin fluorescence and right angle light scatter as measured flow cytometrically. This work in conjunction with the genomic sequence information has better defined the physiological underpinnings of Prochlorococcus LLI-IV ecotype subclades.